A long-term flow cytometry assay to analyze the role of specific genes of Drosophila melanogaster S2 cells in surviving genotoxic stress.
A long-term flow cytometry assay to analyze the role of specific genes of Drosophila melanogaster S2 cells in surviving genotoxic stress.
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长期流式细胞术分析果蝇 S2 细胞特定基因在基因毒性应激中的作用。
DOI：
10.1002/cyto.a.20579
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发表时间：
2008
期刊：
影响因子：
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通讯作者：
Sibon,OdyCM
中科院分区：
文献类型：
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作者：
Yi,Xia;Lemstra,Willy;Vos,MichelJ;Shang,Yongfeng;Kampinga,HarmH;Su,TinTin;Sibon,OdyCM
果蝇S2细胞易于操作和培养，是一种多功能模型系统，用于高通量筛选，例如全基因组siRNA筛选，以查找与应激或治疗抵抗相关的基因，或通过大型化合物库筛选以识别细胞毒素。克隆形成测定被认为是研究特定治疗的细胞毒性或比较不同细胞类型对特定治疗的敏感性的金标准。然而，该测定法不能用于果蝇 S2 细胞，因为它们实际上无法在不同的集落中生长。我们设计了一种新型的基于荧光的流式细胞术测定法来研究 S2 细胞在各种条件下以及特定基因产物存在或特定基因产物下调后的长期增殖。在这里，我们验证了该测定，并使用这种新方法来研究检查点基因grapes/Dchk1和DmChk2在细胞存活反应中的作用。我们的数据表明，Grapes/Dchk1 而不是 DmChk2 是羟基脲生存所必需的。我们的测定将用于研究各种治疗对 S2 细胞的长期影响，并评估其中特定蛋白质的作用。 © 2008 国际细胞计数促进会
DrosophilaS2 cells are easy to manipulate and culture and are a versatile model system for high‐throughput screens such as genome‐wide siRNA screens to find genes involved in stress or therapy resistance or for screening through large compound libraries to identify cytotoxins. Clonogenic assays are considered the gold‐standard to investigate the cytotoxicity of specific treatments or to compare the sensitivity of various cell types for a specific treatment. However, this assay cannot be used forDrosophilaS2 cells as they are virtually unable to grow in distinct colonies. We designed a novel fluorescence‐based flow cytometry assay to study long‐term proliferation of S2 cells under various conditions and in the presence of specific gene products or after downregulation of specific gene products. Here we validate this assay and we used this novel method to investigate the role of checkpoint genesgrapes/Dchk1andDmChk2in cell survival responses. Our data demonstrate that Grapes/Dchk1 but not DmChk2 is required to survive hydroxyurea. Our assay will be of use to investigate the long‐term effects of various treatments in S2 cells and to evaluate the role of specific proteins therein. © 2008 International Society for Advancement of Cytometry